ISSN:1094-9313    

DOI:10.1159/000154865

出版商:S. Karger AG    

年代:1997

数据来源: Karger

ISSN:1094-9313    

DOI:10.1159/000154866

出版商:S. Karger AG    

年代:1997

数据来源: Karger

ISSN:1094-9313    

DOI:10.1159/000154867

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The Kv1.3 potassium channel in T lymphocytes plays a major role in mitogen-induced activation and is widely recognized as a potential therapeutic target for immunosuppressive agents. Availability of structural information on this important protein would greatly facilitate rational drug design. However, determination of the atomic structure of membrane proteins poses overwhelming technical challenges. We have therefore utilized two complementary approaches to probe the architecture of Kv1.3. The first uses structurally defined peptides as molecular probes of the pore region of the channel. These studies reveal the existence of a ∼ 30 Å wide and ∼ 6 Å deep, saucer-shaped external vestibule, at the center of which lies the selectivity filter as a shallow depression. The second strategy exploits a vaccinia viral system to overexpress the Kv1.3 protein in mammalian cells. We have determined the kinetics of Kv1.3 protein synthesis and transport to the membrane in vaccinia-infected cells, and have purified the Kv1.3 protein from these cells to near homogeneity. The purified protein in detergent is tetrameric with x-y dimensions of 65 x

ISSN:1094-9313    

DOI:10.1159/000154868

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

Programmed cell death has been shown to be crucial for the regulation of cellular homeostasis. Ligation of CD95 (Apo-1/ Fas) antigen triggers apoptosis by activating an extensively studied signaling cascade. The present review describes some aspects of CD95/Fas-induced programmed cell death as well as of signaling and deals with the regulation of ion channels during this process. In particular, the n-type K+ channel (Kv1.3) is downregulated upon induction of programmed cell death. The effects of CD95/Fas triggering, ceramide and reactive oxygen species (ROS) on Kv1.3 activity are discussed. The inhibition by the first two factors is mediated by tyrosine phosphorylation of the channel protein. Recent results concerning signaling complexes of tyrosine kinases and potassium channels are described and compared. The possible involvement of other ion channels in apoptosis is also discussed.

ISSN:1094-9313    

DOI:10.1159/000154870

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

Voltage-activated Kv1.3 channels, which are involved in normal functions of human T lymphocytes, contain consensus sites for phosphorylation by PKA, PKC, and CaMKII. Because T-cell functions often depend on second-messenger pathways involving serine/threonine phosphorylation, regulation of Kv1.3 provides a possible means of modulating cell function. This paper presents a brief review of work from my lab on Kv1.3 regulation in human T cells by temperature, PKC, PKA and calmodulin-dependent pathways – results that are discussed in light of potential sites for phosphorylation within the Kv1.3 sequence. Also highlighted are several artefacts that we and others have encountered while studying regulation of this channel, all of which reinforce the caution necessary in interpreting effects of hydrophobic compounds on Kv1.3 current. Poreforming antibiotics commonly used to obtain perforated-patch recordings (nystatin, amphotericin B) directly inhibit Kv1.3 channels, as do several drugs used to activate or inhibit second-messenger pathways (forskolin, trifluoperazine, a synthetic diacylglycerol – 1,2-dioctanoyl-sn-glycerol). Thus, direct actions of such drugs may confound studies of their normal biological ro

ISSN:1094-9313    

DOI:10.1159/000154871

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The type l K+ channel, coded for by the Kv3.1 (synonymous with NGK2, KV4, RKShΠIB, KCNC1) gene, is expressed in murine lymphocytes, central neurons, alveolar epithelium, cardiac muscle, and perhaps myelinated nerve. An abnormally high level of expression occurs in lymphocytes in a number of murine autoimmune diseases and in murine thymocytes and cytotoxic/suppressor lymphocytes. The function of these channels in lymphocytes remains unclear. Type l K+ channels are distinguished by their high threshold voltage for activation, rapid activation and deactivation, slow inactivation with rapid recovery, high sensitivity to block by extracellular but not intracellular TEA+, and insensitivity to peptide toxins that block other K+ channels. Type l channels exhibit strong permeant ion effects on gating. Channel closing is slowed dramatically by Cs+ or Rb+, perhaps due to their binding at a hypothetical site near the external mouth of the channel

ISSN:1094-9313    

DOI:10.1159/000154872

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

In human T lymphocytes patch-clamp experiments have revealed the expression of two major subtypes of SK channels. In excitable cells these channel types play significant roles in shaping excitablility as well as firing patterns especially since their open probability is regulated by the intracellular second messenger Ca2+. In nonexcitable T cells, SK channels could hyperpolarize the membrane during the rise in [Ca2+]i elicited via T-cell receptor stimulation. This hyperpolarization in turn could contribute to enhanced Ca2+ entry by increasing the driving force for Ca2+ to enter the cell [see also Verheugen, this issue] via Icrac· This scenario plays an increasingly significant role in already activated T cells due to the up-regulated number of SK channels in activated human peripheral T lymphocytes. We have used whole-cell recording to determine the electrophysiological properties of current through SKAPA channels in Jurkat T cells and through SKctx channels in activated HPB T lymphocytes. The main similarities between the two channel types are their Ca2+ sensitivity and voltage dependence. The SK channel activity depends steeply on [Ca2+]i, suggesting that multiple Ca2+ must bind to the channel or an associated molecule in order to open the pore. The Ca2+ concentration at which half the channels are activated is Ì° 400-450 nM and the activity of both channel types is voltage-independent. The major difference between the two channel types is their pharmacology regarding peptide toxins like apamin, scyllatoxin, charybdotoxin, as well as other blockers like TEA+. SKcTX and SKapa channels show minor differences in Ba2+ sensitivity, induction time course, selectivity, and single-channel conductance. All these properties together should allow a better distinction between these two different types of SK channels in lymphocytes with the ultimate goal to identify the genes encoding these channe

ISSN:1094-9313    

DOI:10.1159/000154873

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The expression of ion channels in human T lymphocytes changes with the activation state of the cell. In particular the density of a Ca2+-activated K+ [K(Ca)] channel increases dramatically in the first days following activation by T cell receptor (TCR) stimulation. The experiments reviewed in this article show how this channel contributes to an enhancement of the secondary immune response, by mediating positive feedback between Ca2+ influx and membrane hyperpolarization during the Ca2+-response upon TCR restimulation.

ISSN:1094-9313    

DOI:10.1159/000154874

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

In T lymphocytes and most other nonexcitable cells, store-operated channels (SOCs) are activated through an unknown mechanism following depletion of intracellular Ca2+ stores. Several lines of evidence indicate that a specific SOC, known as the Ca2+ release-activated Ca2+ (CRAC) channel, mediates the influx of Ca2+ that occurs in response to stimulation through the T-cell receptor. CRAC channels are characterized by an extremely high Ca2+ selectivity and low unitary conductance and by a sensitivity to positive and negative regulation by calcium. Stimulation of T cells elicits a variety of Ca2+ patterns, including spikes, oscillations, and maintained plateaus. The mechanisms underlying these responses may involve delayed feedback between Ca2+ stores, [Ca2+]i and CRAC channels as well as the uptake and release of Ca2+ by mitochondria. Information may be encoded in the amplitude and duration of Ca2+ signals, as the spike and plateau Ca2+ response differentially activate transcriptional pathways in B cells. CRAC-deficient T-cell mutants and mammalian homologs of the Drosophila trp protein present attractive opportunities for elucidating the molecular basis and consequences of store-operated calcium influx in T cells.

ISSN:1094-9313    

DOI:10.1159/000154875

出版商:S. Karger AG    

年代:1997

数据来源: Karger